Integrins are cell surface heterodimers that are composed of distinct alpha and subunits. Integrins mediate interactions with multiple extracellular and cellular ligands. Integrins are involved in cell migration, cell-cell interaction, development, tumor metastasis, wound healing and in the pathogenesis of many diseases. Integrin/ligand interaction is, therefore, a therapeutic target. We recently identified many structural elements of alpha and beta subunits of integrin involved in ligand binding and regulation. (1) A residue of Beta1 critical for ligand binding, (2) a regulatory epitope of Beta1, which is potentially involved in activation and inhibition, and (3) activation-dependent conformational epitopes of Beta1, which are induced upon activation of Beta1 integrins, (4) critical residues of alpha2 I domain for binding of collagen and echovirus-1, (5) a predicted Beta-turn of the third amino-terminal repeat of the alpha subunit as potentially critical for ligand binding of non-I domain integrins, (6) a putative laminin-5 binding site of alpha3, distinct from that of alpha4. To elucidate more structural basis of ligand/integrin interactions at the molecular level, we propose to (1) identify the regions or residues critical for ligand binding and ligand specificity of the non-I domain integrin alpha subunits using site-directed mutagenesis and recombinant integrin fragments. (2) Identify laminin-5 binding site of alpha3 subunit using site-directed mutagenesis. (3) Identify residues and regions of Beta1 that are critical for ligand binding and regulation to prove the hypothesis that an I domain-like structure is present in Beta1. These projects will facilitate understanding of the ligand/integrin interactions and help to design potential inhibitors/activators that modulate the interaction, which might be useful (e.g., for preventing metastasis, thrombosis, rejection on organ transplantation).